Perturbation of the TCR/CD3 complex with anti-TCR or anti-CD3 antibodies (Ab) is followed by a sequence of biochemical and biological responses similar to those observed subsequent to T-cell interaction with antigen (Ag). We have reported that assembly of cytoskeletal microfilaments occur rapidly in response to Ab perturbation of the CD3 complex. Microfilaments are imortant structural cellular elements and may have a role in internalization, routing, and, possibly, coupling of surface recepotrs with effector mechanisms. Disruption of microfilament assembly with specific inhibitors (cytochalasins) may have functional and biochemical consequences on T cell activation. Cytochalas in pretreatment enhanced hydrolysis of membrane inositol phospholipids (InsPL) induced CD3 perturbatiobn. InsPL hydrolysis by phospholipase C (PLC) generates second messengers which may be involved in signal transduction. Potentiation of InsPL hydrolysis by cytochalasins affected the initial rate and maxima level of the response. Decay rates were unaffected, suggesting that the turn off mechanism is independent of the microskeleton. The effect of cytochalasins did not correlate with the association of the TCR/CD3 complex with detrgent-insoluble cytoskeleton, but was associated with a decreased receptor internalization rate. The relationship between recepotr turnover and internalization, and signal transduction was characterized in detail. Ab immobilized onto solid matrices have been used as an alternative to perturbation of TCR/CD3 with soluble Ab. Conditions and requirements for the interaction of T lymphocytes with immobilized anti-TCR or anti- CD3 Ab have been previously defined. As opposed to soluble Ab, the rigidity of a solid matrix would effectively prevent potentiate InsPL hydrolysis even when receptor internalization is prevented by physical means. This observation indicates that microfilament disruption may favor coupling of the TCR/CD3 complex to the signallying apparatus, and suggests a negative feed-back role for microfilament polymerization in PLC activation.